Multiple biomarker approach in the fiddler crab to assess anthropogenic pollution in tropical coastal environments.

CONTEXT: Fiddler crabs are important to the ecology of estuarine systems around the world, however, few studies have incorporated them as bioindicators. Urias estuary represents one of the most urbanized lagoons in the Gulf of California region and received discharges from different sources: shrimp farm, thermoelectric plant, fish processing plants, and untreated domestic and sewage wastes. OBJECTIVE: Assess the effects on anthropogenic contamination on female fiddler crabs reproduction, survival and genetic stability. METHODS: Exposition of wild crabs from a less impacted (reference) site to naturally contaminated sediments on under controlled laboratory conditions. Reproductive parameters, levels of DNA damage and mortality rates were measured, together with chemical analyses of sediments. RESULTS: The most contaminated sediments corresponded to the site where fish processing plants were located and the integrated biomarker response analysis revealed that the most adverse effects were produced by exposure to sediments from this site; these crabs showed higher mortality (67%) and poorer ovarian development than those crabs exposed to sediments from other sites. CONCLUSIONS: Female crabs under pollution stress are able to trade-off reproduction for survival, and surviving animals were able to restore genetic stability possibly by activating DNA repair mechanisms. Multiple biomarker approach discriminates different coastal contamination scenarios.

The white mullet (Mugil curema) as biological indicator to assess environmental stress in tropical coastal lagoons.

Several coastal lagoons and estuaries in Mexico receive untreated domestic and industrial discharges which contain complex mixtures of contaminants. In order to assess the effects of chemical contamination, we used the White mullet (Mugil curema) as biological indicator. We worked in two estuaries located in Northwest Mexico: Urias (highly polluted) and Teacapan (less polluted, therefore used as reference site). We measured several endpoints at different levels of biological organization: vitellogenin transcription in males as biomarker of estrogenic contamination, as well as reproductive, morphological (deformities), morphometric, and meristic parameters. Total RNA was isolated from the liver, and a partial sequence of the mullet vitellogenin gene was obtained; gene expression was analyzed by quantitative PCR. At the same time, gonad samples were analyzed by histologic techniques to determine sex ratios and the reproductive cycle stage. The reproductive season was detected from February to June in both sites, but the gonadosomatic index was consistently higher in Teacapan. Sex ratios were female-biased in both estuaries, and one intersex gonad and several malformations were found in fish from Urias. Vitellogenin gene transcription in males was detected in both sites, although gene expression was slightly higher in Urias. The results obtained in this study indicate that biological effects of contamination are evident in fish, environmental estrogens may be present in both estuaries, and the white mullet is useful as biological indicator to identify and characterize environmental stressors in tropical coastal ecosystems.

Expression of aromatase in the embryonic brain of the olive ridley sea turtle (Lepidochelys olivacea), and the effect of bisphenol-A in sexually differentiated embryos.

Brain aromatase participates in several biological processes, such as regulation of the reproductive-endocrine axis, memory, stress, sexual differentiation of the nervous system, male sexual behavior, and brain repair. Here we report the isolation and expression of brain aromatase in olive ridley sea turtle (Lepidochelys olivacea) embryos incubated at male- and female-promoting temperatures (MPT and FPT, respectively), at the thermosensitive period (TSP) and the sex-differentiated period. Also, aromatase expression was assessed in differentiated embryos exposed to bisphenol-A (BPA) during the TSP. BPA is a monomer of polycarbonate plastics and is considered an endocrine-disrupting compound. Normal aromatase expression was measured in both forebrain and hindbrain, showing higher expression levels in the forebrain of differentiated embryos at both incubation temperatures. Although no significant differences were detected in the hindbrain, expression was slightly higher at MPT. BPA did not affect aromatase expression neither in forebrains or hindbrains from embryos incubated at MPT, whereas at FPT an inverted U-shape curve was observed in forebrains with significant differences at lower concentrations, whereas in hindbrains a non-significant increment was observed at higher concentrations. Our data indicate that both incubation temperature and developmental stage are critical factors affecting aromatase expression in the forebrain. Because of the timing and location of aromatase expression in the brain, we suggest that brain aromatase may participate in the imprinting of sexual trends related to reproduction and sexual behavior at the onset of sex differentiation, and BPA exposure may impair aromatase function in the female forebrain.

Follicular apoptosis in the mussel (Mytella strigata) as potential indicator of environmental stress in coastal ecosystems.

Follicular apoptosis in the tropical mussel Mytella strigata was assessed in three coastal lagoons located in the southern Gulf of California, Mexico. Mussels were collected from three coastal lagoons associated with different scenarios of anthropogenic stress during one year. The gonad of each mussel was dissected, weighed, and sampled for histology and apoptosis analysis by TUNEL labeling. Two apoptotic indices were used: the apoptotic index of cells (AIC) based on the number of follicular cells in apoptosis in one thousand cells counted per gonad, and the apoptotic index of follicles (AIF) based on the number of follicular cells per follicle per gonad. Both indices showed high association with each other for all developmental stages, although AIF seemed to better discriminate among sites. Higher AIF and AIC were observed at the Urias Estuary (1.6 and 1.5 respectively) ranked as highly polluted, followed by Ensenada del Pabellon (0.82 and 0.95 respectively), ranked as moderately polluted, and the Teacapan Estuary (0.57 and 0.76 respectively) ranked as slightly polluted. Our data indicate that the apoptotic index in tropical mussels could be a useful indicator of environmental stress in coastal ecosystems; however, the ecological relevance of follicular apoptosis in polluted environments needs further investigation.

Heat-shock protein (Hsp70) and cytochrome P-450 (CYP1A) in the white mullet Mugil curema (Pisces:Mugilidae) as biomarkers to assess environmental quality in coastal lagoons.

Biomarkers have been useful tools to monitor some effects of pollution in coastal environments. Hepatic expression of heat-shock protein 70 (Hsp70) and cytochrome P450 1A (CYP1A) were analyzed in white mullet (Mugil curema) by RT-PCR from July, 2005 until July, 2006 in three coastal lagoons located in the southern Gulf of California, Mexico. These three coastal systems receive contaminants derived from local anthropogenic activities. Heat-shock proteins function to maintain protein integrity in the presence of stressors (such as heat or chemicals) and can be used as biomarkers of homeostatic alterations in polluted environments, whereas cytochrome P450 family members participate in steroid hormone synthesis and metabolism, and in xenobiotic transformation as a detoxification mechanism. The expression levels of both genes showed consistency in time and space, and presented a high overall correlation (r = 0.731, P < 0.001). Regardless of a high individual variability, both genes presented higher expression levels in the Urias Estuary (P < 0.001 and P < 0.05 for CYP1A and Hsp70, respectively), which was considered the most polluted among the three systems, especially during the rainy season (summer to fall). Gene expression levels were significantly associated with non-halogenated hydrocarbon concentrations in sediments during the sampling period (r = 0.686, P = 0.019 for CYP1A and r = 0.91, P < 0.001 for Hsp70), suggesting that both genes respond to chemicals in the environment. The results indicate that Mugil curema is a good candidate species to implement biomonitoring programs in tropical coastal environments.

The role of vitellogenin during gestation of Girardinichthys viviparus and Ameca splendens; two goodeid fish with matrotrophic viviparity.

Goodeid fish have matrotrophic viviparity, and unlike lecitotrophic fish, yolk loss forces the female to provide the nutritional requirements for embryonic development. Vitellogenin (VTG) is the yolk precursor protein synthesized in the maternal liver, but there is only circumstantial evidence regarding VTG supply during the ontogenesis of bony fish with matrotrophic viviparity. Therefore, the goal of the present study was to identify and quantify VTG during gestation of the black fin goodeid Girardinichthys viviparus and the butterfly split-fin goodeid Ameca splendens. Females at different gonadic developmental stages were selected in order to evaluate VTG mRNA expression in the maternal liver using RT-PCR; VTG quantification in maternal muscle and liver, as well as in the embryos, was done using ELISA, and immunohistochemical detection of VTG was done in the black fin goodeid. The results suggest that VTG supplies nutrients during embryonic development of both species, which have different life histories. It is possible that the transition from lecitotrophy to matrotrophic viviparity in bony fish with intraluminal gestation involved adaptive transition strategies that included changes in the relationship between oocytes and follicular cells, as well as a gradual loss of VTG synthesis during embryonic development.